PROJECT SUMMARY/ABSTRACT Age is the major risk factor for many diseases including cancer, cardiovascular and neurodegenerative disease. Biogerontology research is well positioned to help prevent or at least postpone these diseases by identifying strategies to delay aging and altering its effects on macromolecular, cellular and extracellular damage so that the degree and type of damage does not reach the threshold required for disease incidence or progression. In Project 1 renewal application, we will study the effects of prolonged fasting cycles (PFC), fasting mimicking diet (FMD) cycles and protein restriction cycles (PRC) and of the GH-IGF-1 axis on the aging of the immune and nervous systems with focus on cellular protection and regeneration/rejuvenation. We propose to improve the fasting-mimicking dietary interventions shown in the previous funding period to promote healthspan and test the effect of bi-monthly cycles of these diets on healthspan in different genetic backgrounds. A central goal of Project 1 will be to identify periodic dietary interventions that extend healthspan without promoting adverse effects at very old ages. A major effort will be devoted to the identification of the molecular mechanisms responsible for the effects of periodic fasting mimicking diets on cellular protection with emphasis on the connection between nutrient signaling, stress resistance transcription factors, and the activation of a protective ketone body-activated alternative metabolic mode. Because of the focus of Projects 2 and 3 on mitochondria and stress resistance, we anticipate synergism between Project 1 and Project 2, which will investigate the effects and mechanisms of action of the DR mimicking mitochondrial peptide humanin (Project 2), and Project 3, which will test the hypothesis that the gas H2S is a central mediator of fasting- depended protection. Finally, based on our preliminary results showing that prolonged fasting increases stem cell-based regeneration in multiple systems, approximately half of the effort of Project 1, will be devoted to understanding the effect of periodic fasting and fasting mimicking diets on the regeneration of hematopoietic and neural stem cells and to the mechanisms underlying these effects. A key question that will be investigated is whether this regeneration results in a functional rejuvenation of the immune and nervous systems. We anticipate that the new insights gained from this project will continue to be translated into clinical trials to identify interventions that are safe and effective in improving human healthspan.